Dr. Hughes' laboratory is focused on developing molecular and cytogenetic technologies useful at the level of a single cell. This includes single cell fluorescent and amplification techniques as well as expression cDNA libraries developed from the single haploid and diploid cell. These techniques have usefulness in a wide array of diagnostic and basic science applications. For example, in tumor biology the ability to analyze a single cell enhances the likelihood of identifying LOH. We have developed a method which is routinely capable of generating a karyotype from a small region of a tumor, even a single cell. The technique (called comparative genome hybridization) was developed initially by others and we have miniaturized the analysis to a single diploid cell. Together with the pathologist, a region as small as one cell can be identified on a frozen or paraffin embedded sample and chromosomal imbalances as well as multiple genetic loci can be studied with high reliability. Tissue culture and conventional cytogenetics are omitted and diagnostic results can be obtained in 1-2 days instead of weeks. More importantly, the data seem be easier to interpret because the sample (one cell) has a homogeneous genotype. Another application is Fetal Cell Sorting From Maternal Blood. In every pregnancy a very few fetal cells cross the placenta-uterine interface and enter the maternal circulation. Technologies now exist to find the 1-in-10 million fetal erthroblasts in a sample of maternal blood. A single fetal cell is identified and selected from the sorted sample and analyzed for its chromosomal karyotype and at multiple, specific genetic loci. Consequently, single gene defects as well as chromosomal aneuploidy can be identified, perhaps someday without the need for invasive testing of the pregnancy (amniocentesis). Preimplantation Genetics is an emerging new field which has directly evolved from our single cell work. Chromosome analysis as well as molecular detection of mutations causing Tay Sachs, Duchenne dystrophy, cystic fibrosis, Huntington disease, and hemophilia A can now be made before pregnancy begins. Single sperm and oocytes are being examined for nucleotide triplet repeat expansions causing myotonic dystrophy, spinocellebellar ataxia and Huntington's, in an attempt to elucidate the basic mechanism(s) behind these "dynamic mutation" disorders. The overall objective is to develop and utilize these new genetic technologies for patient care and basic discovery.